This invention relates to the method for immobilization or fixation of enzymes or enzyme-containing microoganism cells by which enzymes are made insoluble in water and fixed on a vehicle or carrier.
It is well known that enzymes are used as the catalyst that effectively promotes many chemical reactions which cannot be carried out with ordinary chemical catalysts and are indispensable materials in the fields of food industry, pharmaceutical industry and other industries. Heretofore, in most enzymatic reactions, enzymes are used in an aqueous phase and they are discarded together with waste liquids after the reaction. In most cases, it is very difficult from both technical and economic viewpoints to recover enzymes from the reaction mixture for repeated use thereof. Therefore, enzymatic reactions are always carried out batch by batch. So, if a method to insolubilize enzymes and immobilize them in a form in which substrates can be continually contacted with them, enzymes can be used repeatedly and enzymatic reaction processes can be carried out continuously.
So far, it has been attempted to chemically combine enzymes with water-insoluble substances such as synthetic macromolecules so as to fix them. However, the activity of enzymes is very sensitive to change in the molecular structure thereof, and in most cases enzymes are remarkably deactivated by such chemical fixation. Therefore such methods are not successfully employed in practical application. Also it was proposed to make enzymes be adsorbed on an absorbent such as active carbon, active terra alba (white earth), etc. (U.S. Pat. No. 2,717,852, 1955). But in this method, the enzyme adsorbed thereon is easily desorbed as it is repeatedly used for reaction. This method is unpractical because of short life of the enzyme composition. Further, the so-called entrapping or inclusion method is known. For instance, a method which comprises polymerizing a water-soluble monomer suh as acrylamide in a solution containing an enzyme (K. Kawashima et al., Biotech. Bioeng. 16, 609, 1974) and a method which comprises cross-linking polyvinyl alcohol in its aqueous solution containing an enzyme (H. Maeda et al., Biotech, Bioeng. 15 827, 1973) have been proposed. However, these polymers remarkably swell with water and therefore the inclusion mass is in the state of a gel lump that has no pores or voids, and therefore it is necessary to dry up the lump and pulverize it in order to convert it to a form in which the enzyme can contact with a substrate substance. This is a troublesome treatment and involves high possibility of desorption and/or deactivation of the enzyme during such treatment.
We have endeavored to find out a simple and effective method of immobilizing or fixing enzymes securing the long-lasting enzyme activity and now provide a new method of this invention.